Project 1 CVD Case/Control Study 1. Completed phase I MRM on 33 targets on entire CVD sample set We targeted 35 promising proteins from discovery proteomics and assayed them using a more quantitative mass spec method known as Multiple Reaction Monitoring (MRM) in a larger study of 336 CVD cases and 336 controls. Stepwise regression, using all 35 MRM proteins identified a CVD risk multimarker panel that requires further exploration and replication 2. Completed phase II MRM (Depletion-MRM) We targeted 27 additional promising proteins from discovery proteomics and assayed them using depletion-MRM. This technique utilizes an MRM assay employs with depletion of the most abundant proteins prior to running on the Mass Spec to allow recovery of lower abundance biomarker targets. These proteins were measured in the same CVD cases and controls. The data are under analysis and we will build univariate and multivariate panels for CVD risk predictiion. Metabolic Syndrome Study (MetSyn) 1. Completed GCMS data collection on entire MetSyn sample set: GCMS measurements of 149 analytes were completed on the 650 sample set designed for the MetSyn, three axis, factorial study of obesity, dyslipidemia, and dysglycemia. Multiple promising metabolomic markers were identified and were replicated in an external cohort (See #2 below). Results were presented at the 2012 AHA ATVB meeting. We are working with collaborators to understand the underlying biology. 2. Completed Targeted GCMS Replication Study in Bioimage sample set: We took the top 37 GCMS markers from the MeySyn study (see #1 above) and conducted a targeted replication experiement in the Bioimage cohort. The discovery-replication design of this study permitted the identification of metabolites associated with all three metabolic risk factors as well as metabolites specific to individual risk factors. Our findings shed light on the breadth and depth of metabolomic derangements associated with major metabolic risk factors and we are completing a manuscript describing these results. 3. Completed Lipid LC/MS data collection in entire Met Syn sample set Targeted plasma Lipid LC/MS measurements of 154 lipid species analytes were completed on the 650 sample MetSyn study. The data were analyzed and some promising associations were identified for each of the three axes tested. 4. Completed iTRAQ discovery proteomics (Px) in 160 subjects in the MetSyn study iTRAQ proteomic labeling was carried out on 160 samples from the MetSyn study representing a balanced distribution of the sample selection strategy. A total of 860 proteins were detected; 543 were present in 60% or more of the samples. These data were analyzed and we are currently deciding on the next step for these data. Project 2 1. Completed measurements of 4 concentration panels Successful measurement of 34 protein targets across >7500 samples has been completed. The data have been QCd and are in an analysis pipeline to conduct an initial analysis of associations of these proteins with CVD. More targets will be added to the model as the project moves forward and generates more biomarker data. 2. Nominated 16 new biomarker targets We have utilized internal SABRe data to nominate the next 16 targets for assay development. These targets come from various literature and SABRe data mining efforts. Nominations were reviewed by the SABRe Biomarker Nominating Committee. These 16 new targets reflect the first 1/3 of the total that will be measured this year. 3. Posted first 3 data sets to dbGaP The September 12th dbGaP release contains the data for 27 biomarker targets run the in the 7500 Framingham samples. These data are now accessible and open for sharing. The team developed a pipeline for processing these data such that future data sets will be directly deposited as they complete QC processing. 4. Developed an analysis pipeline for QC of data sets and preliminary CVD trait analysis The QC and data posting pipeline is now fully functional; a pipeline for data analysis is needed. We are working towards a process that will allow team statisticians to rapidly analyze each biomarker in association with all cause mortality, CVD, and CHD. Project 3 1. Completion of CVD case/control data analysis including qPCR validation study The data analysis of the CVD case/control study has been informative. We have a two prong approach to this analysis covering the gene expression signatures of CHD where paired analyses were performed with and without covariates and a second analysis aimed at a more systems biology approach looking at coexpression networks and key drivers of CHD. Both of these analytical approaches are nearing publication. 2. Completed the data collection of the Gen III data set The full Third Generation data set was completed and moved into cleaning and QC processing. The cleaning, identify verification and normalization took several months, but the data set is finally ready for analysis in September. 3. Posted the Gen III data set to dbGaP With the final Gen III data set ready for internal distribution, the data set were posted to dbGaP and became available as a public data set with the September 12th, 2012 dbGaP data posting. 4. Initiated vanilla analyses for >12 SHARe Working Groups (117 phenotypes) on the Offspring data set To help jump start the analysis of the Offspring gene expression resource, the NHLBI analysis team conducted an initial expression analysis of 117 Framingham phenotypes. Individual working group meetings were held to share this initial quick peek at the data with the phenotype experts to begin to develop ideas and strategies for the best means to utilize this vast resource. Project 4 1. Completion of the Offspring Cohort miRNA measurements We completed measurement of 350 miRNA across 2500 Offspring Exam 8 participant samples. These data are undergoing QC and data analysis for various CVD and CVD risk factor phenotypes to attempt to understand the regulatory effects of miRNA on CVD and related phenotypes. 2. Initiated miRNA profiling for the Gen III Cohort The identical analytical platform initiated measurement of the Gen III samples (n=3400) in July of 2012 and will complete the measurements in early October, 2012. These data will then be subjected to the same analytical pipeline that was developed for the previous samples. 3. Characterized miRNA signatures of atherosclerotic CVD The CVD case/ control study and the Offspring Exam 8 miRNA data are being statistically analyzed to uncover the relationship of miRNA to atherosclerotic CVD and its related risk factors. This will include adjusting for technical covariates. We will conduct association analyses of miRNA expression with CVD and its major risk factors. 4. Prepared a cleaned and normalized Offspring data set The Offspring data set has been cleaned and normalized to address the many batches needed by the lab to compile this large data set. Many technical factors were considered to adjust the data and provide the most usable data set possible. The final data set was adjusted for 7 technical variables and normalized against 2 repeated housekeeping miRNA. 5. Posted the case/control set and the full Offspring Cohort data to dbGaP The final data sets for the miRNA case/control and the full Offspring data sets were released by dbGaP on September 12, 2012 for public data access.